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Fig. 9 BMP signaling is required between 27 and 37 hpf for runx2b expression and head cartilage development.
(A-E) Cartilage was stained with Alcian Blue in 4 dpf larvae, ventral views are shown, anterior to the left. (A) Type 1 larvae (blue) display a wild-type morphology, all cartilage elements are present and well shapped. (B) Type 2 larvae (pink) lack ceratobranchials and have mis-shaped Meckel’s cartilage, palatoquadrate, ceratohyal and hyosymplectic. (C) Type 3 larvae (green) display a complete absence of viscerocranium and a reduction of the anterior neurocranium. (D,E) Graphs representing the proportions of the three types of larvae after the indicated treatments. (D) Treatment with the BMP inhibitor dorsomorphin (200 μM) most severely affects head cartilage between 29 and 45 hpf; (DMSO) dimethylsulfoxide. (E) Heat shock treatment of (hsp70l:dnBmpr-GFP)w30 transgenic embryos between 27 hpf and 37 hpf most severely affects pharyngeal cartilage development. (F-H) In situ hybridization for runx2b expression at 48 hpf, lateral views, anterior to the left, scale bars 100 μm. (F) Type 1 embryos (orange) have a normal runx2b expression pattern in all pharyngeal cartilage precursor cells, cleithrum and ethmoid plate. (G) Type 2 embryos (purple), compared to type 1 embryos, only express runx2b in cleithrum and weakly in the ethmoid plate. (H) Graph representing the proportions of the two types of larvae after the indicated treatments. Dorsomorphin treatment of wt embryos and heat shock treatment of (hsp70l:dnBmpr-GFP)w30 between 27 hpf and 37 hpf decreases runx2b expression in pharyngeal cartilage, but not in the cleithrum. (DMSO) dimethylsulfoxide, (Tg+) Transgene expressing embryo, (Tg-) Transgene non-expressing siblings, (HS) heat shock. Meckel’s cartilage (m), palatoquadrate (pq), ceratohyal (ch) and hyosymplectic (hs), ceratobranchials 1 to 5 (cb1-5).